Antibiotics, lactam biosynthesis

The (5-lactam antibiotics are now so extensively described that we cannot attempt to summarize the literature. Since our emphasis is on sulfur, we note that the sulfur atoms of the thiazolidine or dihydrothiazine rings derive from a common tripeptide, 8-(L-a-aminoadipyl)-L-cysteinyl-D-valine 1, ACV or Arnstein tripeptide . ACV is converted to a (5-lactam structure, isopenicillin N 2 and thereafter, the two pathways diverge, i.e. to benzylpenicillin 3 or to cephalosporin C 4 (Scheme 1). There have been extensive studies of the genes and enzymes involved in (5-lactam biosynthesis.18,19... 

A Paradkar, SE Jensen, RH Mosher. Comparative genetics and molecular biology of P-lactam biosynthesis. In WR Strohl, ed. Biotechnology of Antibiotics, 2nd ed. New York Marcel Dekker, 1997, pp 241-277. 

Kim MG, Lee SB (1996) Penicillin acylase-catalyzed synthesis of P-lactam antibiotics in water-methanol mixtures effect of cosolvent content and chemical nature of substrate on reaction rates and yields. J Mol Catal B Enzym 1 201-211 Kohsaka M, Domain AL (1976) Conversion of penicDlin N to cephalosporin(s) by ceU-free extracts of Cephalosporium acremonium. Biochem Biophys Res Commun 70(2) 465-473 Koreishi M, Tani K, Ise Y et al. (2007) Enzymatic synthesis of P-lactam antibiotics and /V-fatty-acylated amino compounds by the acyl transfer reaction catalyzed by peniciUin V acylase from Streptomyces mobaraensis. Biosci Biotechnol Biochem 71(6) 1582-1586 Kupka JY, Shen, YQ, Wolfe S et al. (1983) Partial purification and properties of the alpha-ketoglutarate-linked ring expansion enzyme of beta-lactam biosynthesis of Cephalosporium acremonium. FEMS Microbiol Lett 16 1-6... 

P-Lactam Antibiotics.—The biosynthesis of /3-lactam antibiotics has been reviewed. ... 

These two examples illustrate that both precursor flux, in the case of p-lactam biosynthesis, and antibiotic biosynthetic enzyme levels, in the case of tyiosin, can con-... 

Paradkar, A., Jensen, S.E. and Mosher, R.H. (1997) Comparative genetics and molecular biology of -lactam biosynthesis. In Strohl, W.R. (ed.). Biotechnology of Antibiotics. Marcel Dekker, New York, pp. 241-277. 

Antibiotics have a wide diversity of chemical stmctures and range ia molecular weight from neat 100 to over 13,000. Most of the antibiotics fall iato broad stmcture families. Because of the wide diversity and complexity of chemical stmctures, a chemical classification scheme for all antibiotics has been difficult. The most comprehensive scheme may be found ia reference 12. Another method of classifyiag antibiotics is by mechanism of action (5). However, the modes of action of many antibiotics are stiU unknown and some have mixed modes of action. Usually within a stmcture family, the general mechanism of action is the same. For example, of the 3-lactams having antibacterial activity, all appear to inhibit bacterial cell wall biosynthesis. 

P-Lactams. AH 3-lactams are chemically characterized by having a 3-lactam ring. Substmcture groups are the penicillins, cephalosporias, carbapenems, monobactams, nocardicias, and clavulanic acid. Commercially this family is the most important group of antibiotics used to control bacterial infections. The 3-lactams act by inhibition of bacterial cell wall biosynthesis. 

Occurrence, Fermentation, and Biosynthesis. Although a large number of Streptomjces species have been shown to produce carbapenems, only S. cattkja (2) and S. penemfaciens (11) have been reported to give thienamycin (2). Generally the antibiotics occur as a mixture of analogues or isomers and are often co-produced with penicillin N and cephamycin C. Yields are low compared to other P-lactams produced by streptomycetes, and titres are of the order of 1—20 p-g sohdusmL despite, in many cases, a great deal of effort on the optimization of the media and fermentation conditions. The rather poor stabiUty of the compounds also contributes to a low recovery in the isolation procedures. The fermentation and isolation processes for thienamycin and the olivanic acids has been reviewed in some detail (12). 

P-lactam antibiotics, exert thek antibacterial effect by interfering with the synthesis of the bacterial cell wall. These antibiotics tend to be "kreversible" inhibitors of cell wall biosynthesis and they are usually bactericidal at concentrations close to thek bacteriostatic levels. Cephalospotins are widely used for treating bacterial infections. They are highly effective antibiotics and have low toxicity. 

Now that we have provided you with an overview of the history of penicillin production, we will examine some more details of the biotransformation of -lactams. We will briefly outline the normal biosynthesis pathways that lead to their production and then consider how these products may be diversified in vitro to give a wider range of valuable compounds. We begin by briefly explaining how the fi-lactam antibiotics are effective as therapeutic agents. 

In this chapter, by using the examples of -lactams we have briefly examined how microbial cultures may be used to produce sufficient antibiotics to meet market demands. We have also explained how enzymes (or cells) may be used to biotransform, and thereby diversify, antibiotics. By outlining the history of penicillin production, we explained how analysis and manipulation of culture regimes may be used to enhance the yields of antibiotics (and other secondary products). These studies led to die concept of directed biosynthesis by precursor feeding. 

The antibiotic activity of certain (3-lactams depends largely on their interaction with two different groups of bacterial enzymes. (3-Lactams, like the penicillins and cephalosporins, inhibit the DD-peptidases/transpeptidases that are responsible for the final step of bacterial cell wall biosynthesis.63 Unfortunately, they are themselves destroyed by the [3-lactamases,64 which thereby provide much of the resistance to these antibiotics. Class A, C, and D [3-lactamases and DD-peptidases all have a conserved serine residue in the active site whose hydroxyl group is the primary nucleophile that attacks the substrate carbonyl. Catalysis in both cases involves a double-displacement reaction with the transient formation of an acyl-enzyme intermediate. The major distinction between [3-lactamases and their evolutionary parents the DD-peptidase residues is the lifetime of the acyl-enzyme it is short in (3-lactamases and long in the DD-peptidases.65-67... 

The pathway from simple molecules to the peptidoglycan of the bacterial cell wall is lengthy and complex. Many of the details are well known but need not concern us here. Suffice it to say that long carbohydrate chains are synthesized, subsequently decorated with shorter amino acid chains, and these are finally cross-linked to provide a strong strnctnre. It is this final cross-linking step that is inhibited by the p-lactam antibiotics. The consequence is that cell wall biosynthesis cannot be completed and cell death ensnes. Again, the mammalian host carries out no similar reactions so that similar consequences do not ensne for the host orgaiusm. 

Demain AL, Vaishnav P. (2002) Regulation of (l-lactam antibiotic biosynthesis by carbon sources. Chim Oggi/Chem Today 20(11-12) 46-51. 

As already noted, beta-lactam antibiotics interfere with biosynthesis of the primary component of cell membranes—pepfidoglycan. Because of the fact that this process does not take place in human and other mammalian cells, beta-lactam antibiotics are relatively non-toxic to humans. 

During the biosynthesis of the cell wall, the muropeptide is formed from acetylmuramyl-pentapeptide, which terminates in a D-alanyl-D-alanine. The synthesis of this precursor is inhibited by the antibiotic cycloserine (9.36), a compound produced by many Streptomyces fungi but which is not used clinically. During the crosslinking of the pen-tapeptide precursor, the terminal fifth alanine must be split off by a transpeptidase enzyme. This last reaction in cell wall synthesis is inhibited by the p-lactam antibiotics. 

The biosynthesis of cell wall peptidoglycan, showing the sites of action of five antibiotics (shaded bars 1 = fosfomycin, 2 = cycloserine, 3 = bacitracin, 4 = vancomycin, 5 = 3-lactam antibiotics). Bactoprenol (BP) is the lipid membrane carrier that transports building blocks across the cytoplasmic membrane M, N-acetylmuramic acid Glc, glucose NAcGIc or G, /V-acetylglucosamine. 

Intrinsic Activity. y3-Lactam antibiotics affect sensitive bacteria by inhibiting late stages in tire biosynthesis of their cell wall peptidoglycan. 

Both the cephalosporins and the penicillins owe their antibacterial action to their ability to block bacterial cell-wall biosynthesis. Cephalosporin C is less active than the penicillins, but is less susceptible to enzymatic destruction by /3-lactamases, which are enzymes that cleave the lactam ring. In fact, the so-called resistance of staph bacteria to penicillins is attributed to the propagation of strains that produce /3-lactamase. Numerous semisynthetic penicillins and cephalosporins have been made in the hope of finding new broad-spectrum antibiotics with high activity but with greater /3-lactam stability. Several of these are in clinical use. 

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